LOCUS       sequence-224981-        5389 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express myc-Rheb-Omp25.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5389)
  AUTHORS   Lawrence RE, Cho KF, Rappold R, Thrun A, Tofaute M, Kim DJ, 
            Moldavski O, Hurley JH, Zoncu R
  TITLE     A nutrient-induced affinity switch controls mTORC1 activation by its
            Rag GTPase-Ragulator lysosomal scaffold.
  JOURNAL   Nat Cell Biol. 2018 Sep;20(9):1052-1063. doi: 
            10.1038/s41556-018-0148-6. Epub 2018 Jul 30.
  PUBMED    30061680
REFERENCE   2  (bases 1 to 5389)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41556-018-0148-6"; journalName: "Nat Cell Biol"; date: 
            "2018-09"; volume: "20"; issue: "9"; pages: "1052-1063"
COMMENT     Sequence Label: pRK5 Myc Rheb Omp25
FEATURES             Location/Qualifiers
     source          1..5389
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     548..568
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     594..618
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        828..846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     828..845
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     859..878
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      925..934
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             934..963
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     regulatory      1689..1698
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    1702..1836
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1739..1758)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1793..1812
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        1905..2262
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1905..2096
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1928..1948)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2113..2248
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2282..2299)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2282..2298)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2291..2313)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2511..2966
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2598..2617)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2808..2829
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2983..3002)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3102..3124
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3162..3180)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3248..3352
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3353..3421,3422..4213)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3571..3590)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4384..4972
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4873..4892
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5126..5143
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5260..5281
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5296..5301,5302..5319,5320..5326)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5334..5350
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     5339..5361
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5358..5374
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5358..5374
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
      601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga
      661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt
      721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta
      781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
      841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
      901 acctcggttc tatcgattga attcgccacc atggagcaaa agctcatttc tgaggaagat
      961 ctcatgccgc agtccaagtc ccggaagatc gcgatcctgg gctaccggtc tgtggggaaa
     1021 tcctcattga cgattcaatt tgttgaaggc caatttgtgg actcctacga tccaaccata
     1081 gaaaacactt ttacaaagtt gatcacagta aatggacaag aatatcatct tcaacttgta
     1141 gacacagccg ggcaagatga atattctatc tttcctcaga catactccat agatattaat
     1201 ggctatattc ttgtgtattc tgttacatca atcaaaagtt ttgaagtgat taaagttatc
     1261 catggcaaat tgttggatat ggtggggaaa gtacaaatac ctattatgtt ggttgggaat
     1321 aagaaagacc tgcatatgga aagggtgatc agttatgaag aagggaaagc tttggcagaa
     1381 tcttggaatg cagctttttt ggaatcttct gctaaagaaa atcagactgc tgtggatgtt
     1441 tttcgaagga taattttgga ggcagaaaaa atggacgggg cagcttcaca aggcaagtct
     1501 tcatgctcgg tgatggcggc cgcgggcgga ggtggatctg gcggaggtgg atcggttcat
     1561 cgaggcgacg gagagccgag tggagttcct gtagctgtgg tgctgctgcc agtgtttgcc
     1621 cttaccctgg tagcagtttg ggccttcgtg agataccgaa agcagctctg ataaggatcc
     1681 ccagcttggc cgccatggcc caacttgttt attgcagctt ataatggtta caaataaagc
     1741 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     1801 tccaaactca tcaatgtatc ttatcatgtc tggatcggga attaattcgg cgcagcacca
     1861 tggcctgaaa taacctctga aagaggaact tggttaggta ccttctgagg cggaaagaac
     1921 cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     1981 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     2041 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     2101 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     2161 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     2221 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctgtt aacagcttgg
     2281 cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc
     2341 gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc
     2401 gcccttccca acagttgcgc agcctgaatg gcgaatggcg cctgatgcgg tattttctcc
     2461 ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg
     2521 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     2581 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     2641 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     2701 gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg
     2761 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     2821 ccaaactgga acaacactca accctatctc gggctattct tttgatttat aagggatttt
     2881 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
     2941 taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc
     3001 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     3061 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     3121 gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt
     3181 tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg
     3241 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     3301 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     3361 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     3421 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     3481 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     3541 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     3601 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     3661 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     3721 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     3781 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     3841 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     3901 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     3961 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     4021 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     4081 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     4141 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     4201 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     4261 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     4321 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     4381 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     4441 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     4501 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     4561 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     4621 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     4681 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     4741 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     4801 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     4861 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     4921 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     4981 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     5041 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     5101 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
     5161 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag
     5221 gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca
     5281 ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag
     5341 cggataacaa tttcacacag gaaacagcta tgacatgatt acgaattaa
//
